The purpose of this project is to provide new knowledge of the auditory mechanisms of the inner ear. We have continued to study the distribution in the organ of hearing of neurotransmitter candidates and associated enzymes, using small mammals (guinea pigs, rats and mice). We have used polyclonal antisera and monoclonal antibodies, studying the distribution of immunoreactivity through light and electron microscopy in immunohistochemical studies. High performance liquid chromatography (HPLC), radio immunoassays (RIA), and receptor binding experiments were used in biochemical studies of opioid peptides in the organ of hearing. Both the normal and the de-efferented organ of hearing were studied. Several opioid peptides were found in the organ of hearing of the guinea pig. Its lateral system of efferent neurons specifically contains methionine enkephalin, while the medial system contains other peptide(s). In cells of origin of the lateral system of efferents enkephalin-like and choline acetyltransferase-like immunoreactivities are co-contained; such co-containment has previously not been demonstrated in nerve cells. Glutamic acid decarboxylase (GAD)-like immunoreactivity is present in a subpopulation of efferent neurons in the organ of hearing, in both the lateral and the medial system of efferents. This indicates that the present dichotomy of these neurons may need to be modified. We have also used antiserum against neuron-specific enolase (NSE) in a light microscopy study of the hearing organ. We found NSE-like immunoreactivity in inner hair cells but not in outer hair cells, indicating that there may be a major difference in demands on metabolism between the two types of hair cells. The likely presence of NSE in a major type sensory cell has previously not been demonstrated.